Dysregulated wound healing in the cornea can lead to corneal scars and corneal haze resulting in a loss of visual acuity. While many elements of corneal wound healing are well characterized, the biophysical attributes of the wound space and the impact of these on cellular activities critical to wound healing have not been adequately investigated. Along with soluble signaling molecules present in the extracellular milieu, biophysical cues are emerging as key factors in determining cell behaviors. We propose to quantitatively analyze the alterations in stromal topography and compliance throughout the wound healing process. Our hypothesis is that the biophysical cues in the wound space alter throughout wound healing and that these directly influence the transition of keratocytes to myofibroblasts in the corneal stromal layer. The myofibroblasts, in turn, would alter the compliance of the wound microenvironment through active contraction. Ultimately, the change in the biophysical cues in the wound bed would activate apoptotic pathways, promoting removal of the myofibroblasts and an optically clear cornea would return. Using the measured alterations in the biophysical environment as a guide, we will fabricate hydrogel substrates with similar biophysical properties (of compliance and topography) to determine the cellular consequences of these alterations on myofibroblast behavior in vitro. The information that we obtain about the cellular behaviors and gene and protein alterations could enable the therapeutic targeting of biophysical cues during wound healing. Attributes of the wound bed could be directly modulated, such as the contractile elements of the corneal stromal cells, to improve wound healing outcomes. PUBLIC HEALTH RELEVANCE: Wound healing in the cornea can lead to corneal scars and can lead to impairment or loss of vision. While many elements of corneal wound healing are well characterized, biophysical attributes of the wound space have not been adequately investigated. Along with soluble signaling molecules present in the extracellular milieu, biophysical cues are emerging as key regulators of cell behaviors. This proposal would determine the biophysical cues (specifically topography and compliance) present throughout corneal sromal wound healing. Our hypothesis is that the biophysical attributes of the wound space changes throughout the healing process and that the transition of phenotypes of the cells in the corneal stromal layer is modulated by alterations in the biophysical cues of the wound space. In addition, the myofibroblast, a central player in normal and dysregulated wound healing will modify the compliance of the wound microenvironment and thereby assist in wound repair, but also in ultimately triggering signals necessary to prevent corneal haziness. If proven true, our hypothesis would indicate that direct modulation of the biophysical properties and contractile elements of the corneal stromal cells could improve wound healing outcomes.